SUMMARY

The coordination of chloroplast and nuclear genome status is critical for plant cell function. Here, we report that Arabidopsis CHLOROPLAST AND NUCLEUS DUAL-LOCALIZED PROTEIN 1 (CND1) maintains genome stability in the chloroplast and the nucleus. CND1 localizes to both compartments, and complete loss of CND1 results in embryo lethality. Partial loss of CND1 disturbs nuclear cell-cycle progression and photosynthetic activity. CND1 binds to nuclear pre-replication complexes and DNA replication origins and regulates nuclear genome stability. In chloroplasts, CND1 interacts with and facilitates binding of the regulator of chloroplast genome stability WHY1 to chloroplast DNA. The defects in nuclear cell-cycle progression and photosynthesis of cnd1 mutants are respectively rescued by compartment-restricted CND1 localization. Light promotes the association of CND1 with HSP90 and its import into chloroplasts. This study provides a paradigm of the convergence of genome status across organelles to coordinately regulate cell cycle to control plant growth and development.

INTRODUCTION

Chloroplasts in plants and algae have an endosymbiotic origin: over 2 billion years ago, a photosynthetic cyanobacterium-like organism was stably engulfed by a heterotrophic unicellular eukaryote.1 This initial event was followed over the course of evolution by massive gene transfer from the green prokaryote genome to the host-cell nucleus, leading to the complete dependency of the endosymbiont on its surrounding cells and the generation of a novel type of organism, photosynthetic eukaryotes.2 The development of higher plants requires the tight coordination of chloroplast development from undifferentiated proplastids in meristematic cells or from etioplasts, which differentiate from proplastids when seeds germinate in darkness. In addition to being the site of photosynthesis in leaf mesophyll cells, chloroplasts house essential metabolic pathways and contribute to storage and pigmentation in specialized cells.3

Eukaryote genomes are organized into multiple chromosomes whose replication initiates from numerous origins and is strictly regulated.4 To maintain genome integrity and stability, genomic DNA must be replicated completely and accurately, but also only once during the S phase of the cell cycle. The formation of a pre-replication complex (pre-RC) is required for the subsequent ordered assembly of replication factors such as the Origin Recognition Complex (Orc), Cell Division Cycle6 (Cdc6) in budding yeast (Saccharomyces cerevisiae) or its ortholog CDC18 in fission yeast (Schizosaccharomyces pombe), chromatin licensing and DNA replication factor 1 (CDT1), and Mini Chromosome Maintenance (MCM) proteins (MCM2–7).5 MCM proteins are essential replication initiation factors that were originally identified as proteins required for minichromosome maintenance, plasmid replication, or cell-cycle progression in yeast.6 Indeed, cell-cycle checkpoints are crucial for genome integrity, as they stop cell-cycle progression upon DNA damage until repairs have been completed. Such mechanisms are particularly important in plants, which endure daily exposure to DNA-damaging agents such as light and reactive oxygen species (ROS).7

RESULTS

CND1 is a nucleus and chloroplast dual-localized protein
To understand the functional coordination between the chloroplast and nucleus, we systematically screened proteins with predicted chloroplast and nucleus dual localization by combining bioinformatics and proteomics analyses. We focus here on one such putative chloroplast and nucleus dual-localized protein, encoded by the nuclear gene At1g32730 that we named CHLOROPLAST AND NUCLEUS DUAL-LOCALIZED PROTEIN 1 (CND1). CND1 contains four predicted functional domains: two targeting domains, consisting of an N-terminal chloroplast transit peptide (cTP) (amino acids [aa] 1–65) based on TargetP prediction27 and a nuclear localization sequence (NLS) containing the amino acid motif KRKK28; a zinc finger domain (ZF); and the domain of unknown function DUF702 (Figure 1A ).

Abstract

     Agriculture plays a significant part in India due to their population growth and increased food demands. Hence, there is a need to enhance the yield of crop. One of these important effects on low crop yields is diseases caused by bacteria, fungi and viruses. This can be prevented and handled by means of applying plant disease detection approaches. Machine learning techniques will be employed in the process of disease identification on plants as it mostly applies information themselves and offers fabulous techniques for detection of plant diseases. Methods based on Machine learning can be employed for the identification of diseases because it mainly applies on data superiority outcomes for specified task. In this approach, a comprehensive review has been made on the various techniques employed in plant disease detection using artificial intelligence (AI) based machine learning and deep learning techniques. Likewise, deep learning has also gained a great deal of significance in offering better performance outcome for detecting plant disease in the computer vision field. The deep learning advancements were employed to a range of domains that leads to great attainment in the machine learning and computer vision areas. The comparative study is made in terms of machine and deep learning techniques and their performance and usage in various research papers is related to show the effectiveness of deep learning model over machine learning model. In order to prevent major crop losses, the deep learning technique can be used to detect the leaf diseases from captured images.

Introduction

     The advancements of IoT, AI and the Unmanned Aerial Vehicles are integrated together to provide the support to agricultural fields to detect the plant leaf diseases and report that properly to the respective individuals with proper accuracy ranges. In this modern civilization, nobody is interested in farming and agriculture due to the hurdles the farmers are facing every day. So, that all young generation people are switch over their residence to modern cities to lead a safe life and avoid such agriculture field hurdles. The issue of the proficient plant diseases protection is closely linked to viable change in climate and agriculture [13]. Studies show that climate change may vary pathogenic stages and rates; host resistance may also be altered, leading to physiological variations in host-pathogen co-operations [23]. The actuality that nowadays, diseases more freely transferred around the globe than ever before complicates the situation. New diseases may occur where they have not been identified previously and, inherently, where local expertise to combat them is not available [27] (see Table 1).

Conclusion

     An extensive research study is conceded out on various kinds of machine and deep learning techniques for plant disease recognition and classification. After this, other techniques of classification in machine learning might be employed for may be used for plants disease detection and in the intellect of aiding the farmers an automatic disease detection of all kinds of disease in the crop that were to be detected. This analysis discusses various approaches of DL for the plant diseases detection. Furthermore, several techniques/mappings were summarized for recognizing the disease symptoms. Here the development of deep learning technologies in recent years for the identification of plant leaf diseases. We anticipate that this work will be a useful tool for scientists looking into plant disease detection. Also, a comparative study is also made between machine and deep learning techniques. Though a great deal of noteworthy progress was noticed in recent years, there were still some research gaps that should be addressed and to implement effective techniques for plant disease detection.

Highlights

Abstract

Graphical abstract

Keywords

1. Introduction

2. Literature search strategy & statistics

3. Historical framework: early hormetic findings

4. Radiation – UV and ionizing

5. Pollution and pollen

6. Polyamines

7. Discussion

8. Conclusion

CRediT authorship contribution statement

Declaration of competing interest

Acknowledgement

Appendix A. Supplementary data

References

Abstract

This paper evaluated the occurrence of hormetic dose responses in pollen reported over the past eight decades. Hormetic doses responses were induced by a wide range of chemical and physical agents in 34 plant species for pollen germination and pollen tube growth/elongation. Agents inducing such hormetic dose/concentration responses in pollen included nutrients, growth-promoting agents, plant and animal hormones, toxic substances, including heavy metals such as cadmium, gaseous pollutants such as ozone, as well as ionizing and non-ionizing radiation. This paper provides further evidence for the broad generality of the hormesis dose response, supporting substantial prior findings that the hormetic response is independent of biological model, inducing agent, and endpoints measured. Given the widespread potential of inducing hormetic dose responses in pollen, these findings indicate the need to explore their emerging biological, ecological, agricultural, economic and public health implications.

1. INTRODUCTION

In flowering plants, the reproduction process is mediated by the pollen tube. This tubular structure provides the vehicle by which two immobile sperm cells are transported to the ovule, the location of two female reproduction cells (Sprunck, 2010; Dresselhaus and Franklin-Tong, 2013; Vogler et al., 2014). The pollen tube develops only after the pollen grain is transported, attaches, and then rehydrates on the receptive papillae that reside on the top of the stigma. After this process, the pollen grain becomes activated to form the so-called pollen tube, which may be characterized as a tubular protrusion that is chemically directed via complex physical and sensory processes through the pistil, eventually reaching the female gametophyte. At this point in the process, the male reproductive cells finally arrive at the target destination and initiate the process of fertilization (Palanivelu and Tsukamoto, 2012). Beyond their essential role in fertilization, pollen tubes are widely used to assess biological polarity, with broad applications to other cell types and biological processes. Of considerable theoretical and practical significance is that the pollen grain displays a self-organizing system that permits pollen to germinate, producing pollen tubes in vitro within an external signal mediated framework. The pollen germination rates, pollen tube growth rates, and final tube length are typically lower in in vitro studies than those observed in whole plants, most likely due to the absence of female factors that affect pollen germination, tube growth and growth directionality (Johnson and Presse, 2002; Chae and Lord, 2011). The selection of plants for pollen studies is affected by many species-specific processes as there are many factors that affect pollen tube growth, including the morphology of the reproductive tract (e.g. the stigma, style, the septum epidermis, and the funiculus). The nutrient containing extracellular matrix (ECM) contains a nutrient mixture that is important in enhancing pollen tube elongation Journal Pre-proof Journal Pre-proof 4 (Johnson and Presse, 2002; Chae and Lord, 2011). However, many studies employ bi-cellular pollen from species such as tobacco, which yields reliable findings (Volger et al., 2014).

Highlights

Abstract

Keywords

Introduction

Material and methods

Results

Discussion

Conclusions

Declaration of competing interest

Acknowledgments

Appendix A. Supplementary data

References

Abstract

Chloroxylon swietenia DC. (Rutaceae) is widely used in folk medicine and provides a commercial timber. In the present study, phenological events such as flowering and fruiting phenology, pollinator visitation, anthesis, pollen viability, and pollen tube germination tests were analyzed in 11 populations, in Tamil Nadu, India. The number of flowers was recorded as 82.6 ± 1.4 to 162.2 ± 4.2 during flower bud formation and reduced to 25.4 ± 0.5 to 61.8 ± 1.0 at maturity. The number of fruits per twig was 21.0 ± 0.8 to 47.2 ± 0.7 in initial stages and slightly reduced to 16.2 ± 1.8 to 34.6 ± 1.4 at maturation. Sixty-two species of insects were visiting the flowers with bees/ants as effective pollinators. The 2,5-diphenyltetrazolium bromide assay revealed more viable pollen grains in five populations, and fluorescence dye assay revealed maximum viable pollen grains in six populations. In the pollen tube germination test, the 60% sucrose concentration provides significant results, with a high number of germinated pollen grains. Principal component analysis of flowering and fruiting phenology, floral visitors, and pollen viability showed 79.5%, 79.3%, 56.7%, and 41.7% for the first factor, respectively. Our results indicated that C. swietenia depends heavily on pollinators for its reproduction. Hence, there is a need to expand this study to related species which are having agronomic, pharmacological, and economic potential.

Introduction

Anthropogenic growth in Earth’s climate is affecting many aspects of ecological systems (Traill et al 2010), including seasonal timing of biological events, which includes leaf, flowering, and fruiting phenology. Association between phenology and reproductive events in plants with diverse climatic conditions has been well understood; mainly, flowering phenology has a major impact on plant reproductive biology, and its success concerns evolutionary and ecological consequences (Tadey 2020). Flowering phenology is driven by climatic conditions, and this enables the plants to bloom when the surrounding climatic features are most suitable, ensuring effective sexual reproduction (Theobald et al 2017). The importance of phenology to the science of global change has been progressively renowned during the last few decades (Schwartz 1998; Fitter and Fitter 2002; Tandon et al 2003; Selwyn and Parthasarathy 2006; Singhal et al 2011; Tadey 2020).

Pollination biology is a prime process in plant reproductive achievement, frequency, and identity of floral visitors, the ability for autonomous self-pollination, and the magnitude of pollen limitation, in which pollinators transmit vital ecosystem facilities to diverse ecosystems such as natural ecosystems and agroecosystems (Tandon et al 2003; Aizen and Harder 2007; Tadey 2015; Richardson et al 2017; Figueiredo et al 2020; Impe et al 2020). Diversity of wildflowers is maintained by pollination, and it plays an important role in resolving floral resource–related problems such as herbivores and seed predators (Potts et al 2006). Most of the plants depend on animal pollination for their sexual reproduction via insects, birds, bats, and so on, among which insects play a vital role in pollination. It is well accepted that diversity of pollinators or the number of interactions established by a specific plant is defined by different features, for example, resource availability (quantity of nectar or pollen), morphological traits such as color, size, and shape of flowers, and flowering time (Bosch et al 1997); significant association was found between flower color and flowering phenology (Warren and Billington 2005).

1- Introduction

2- Methodology

3- Results

4- Discussion

5- Conclusion

References

Abstract

The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) system enables precision editing of the genome of many plant species. Developing robust gene editing tools in banana and plantain would pave the way for the improvement of these crops. Here, we developed efficient CRISPR/Cas9 genome editing protocol for banana and plantain using multiple gRNAs targeting phytoene desaturase (PDS) gene. CRISPR/Cas9 construct containing two gRNAs was delivered into embryogenic cell suspension cultures of banana cultivar ‘Sukali Ndiizi’ and plantain cultivar ‘Gonja Manjaya’ by Agrobacterium-mediated transformation. The regenerated genome-edited events of ‘Sukali Ndiizi’ and ‘Gonja Manjaya’ showed albino and variegated phenotypes indicating mutations at the targeted sites disrupting the function of PDS gene. The majority of events (52/77 for ‘Sukali Ndiizi’ and 16/17 for ‘Gonja Manjaya’) were albino. Sequencing of the target sites confirmed presence of indels in all the 18 events sequenced demonstrating mutation efficiency of 100 % in both cultivars. The majority of events (6/8) of ‘Gonja Manjaya’ showed indels at both the target sites of PDS gene, however only 2/10 events of ‘Sukali Ndiizi’ showed indels at both sites, with one event (S24) having a knockout of large fragment (723 bp) indicating that both gRNAs were effective. Several of the albino events of both ‘Sukali Ndiizi’ and ‘Gonja Manjaya’ showed homozygous mutations. Further sequencing of four potential off-target sites in five events showed no mutations indicating CRISPR/Cas9 based editing in banana and plantain is targeted and precise with a very low probability of off-target sites. This study could provide a methodological framework for single or multiple knockouts in banana and plantain.

Introduction

Genome editing technologies using specific nucleases have been developed as effective genetic engineering methods for targeted mutations at specific sites in the genome of an organism [1]. CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 (CRISPR associated protein 9) based genome editing is the emerging most powerful tool with ability to create targeted mutations in the genome of crops and to understand the functional biology. These nucleases respond in a target-specific manner and induce a double-strand break (DSB) in the gene sequence [2]. The DSB is repaired either by homologous recombination (HR) or errorprone non-homologous end-joining (NHEJ) mechanisms, which may lead to disruption of gene function [3]. CRISPR/Cas9 system has emerged as a potent genome editing tool due to its simplicity, design flexibility, and high efficiency, and has been applied successfully in several plant species [4]. It is based on the type II CRISPR/Cas immune system of bacteria that protects against invading DNA viruses and/or plasmids. The CRISPR/Cas9 molecular immunity system is made up of the Cas9 endonuclease from Streptococcus pyogenes and a single guide RNA (sgRNA). The sgRNA directs the Cas9 endonuclease to a target sequence complementary to the 20 nucleotides preceding the protospacer-adjacent motif (PAM) NGG or NAG, which is required for Cas9 activity [5-6]. Multiplexing of CRISPR/Cas9 system by integrating two or more gRNAs, which are distance apart, is useful in generating large deletions in plants [7].

Tissue-Specific Gene Knockdown

Advantages and Applications of Tissue-Specific Gene Knockdown

Limitations of Tissue-Specific Gene Knockdown

References

The habituation of cultured cells to cellulose biosynthesis inhibitors such as dichlobenil (dichlorobenzonitrile, DCB) has proven a valuable tool to elucidate the mechanisms involved in plant cell wall structural plasticity. Our group has demonstrated that maize cells cope with DCB through a modified cell wall in which cellulose is replaced by a more extensive network of highly cross-linked feruloylated arabinoxylans. In order to gain further insight into the contribution of phenolics to the early remodelling of cellulose-deficient cell walls, a comparative HPLC-PAD analysis was carried out of hydroxycinnamates esterified into nascent and cell wall polysaccharides obtained from non-habituated (NH) and habituated to low DCB concentrations (1.5 μM; H) maize suspensioncultured cells. Incipient DCB-habituated cell walls showed significantly higher levels of esterified ferulic acid and p-coumaric acid throughout the culture cycle. In terms of cell wall fortification, ferulic acid is associated to arabinoxylan crosslinking whereas the increase of p-coumaric suggests an early lignification response. As expected, the level of hydroxycinnamates esterified into nascent polysaccharides was also higher in DCB-habituated cells indicating an overexpression of phenylpropanoid pathway. Due to their key role in cell wall strengthening, special attention was paid into the dimerization pattern of ferulic acid. A quantitative comparison of diferulate dehydrodimers (DFAs) between cell lines and cell compartments revealed that an extra dimerization took place in H cells when both nascent and mature cell wall polysaccharides were analysed. In addition, qualitative differences in the ferulic acid coupling pattern were detected in H cells, allowing us to suggest that 8-O-4′-DFA and 8-5′-DFA featured the ferulic acid dimerization when it occurred in the protoplasmic and cell wall fractions respectively. Both qualitative and quantitative differences in the phenolic profile between NH and H cells point to a regioselectivity in the ferulate dehydrodimerization.

Introduction

Primary cell walls of commelinoid monocots (also known as type II cell walls) such as maize are composed of a framework of cellulose microfibrils embedded in a hemicellulosic matrix of (glucurono)arabinoxylans and a smaller amount of pectins and glycoproteins (Carpita and Gibeaut, 1993). These cell walls are also characterised by the presence of cell wall phenolics, mainly the hydroxycinnamates, ferulic (FA) and p-coumaric (CA) acids, which are ester-linked to α–L-arabinosyl residues of (glucurono)arabinoxylans (Smith and Hartley, 1983; Wende and Fry, 1997). Although phenolics are minor components of the primary cell wall (~1% of cell wall dry weight), their contribution to cell wall assembly seems to be crucial. In vitro experiments have demonstrated that hydroxycinnamates are susceptible to oxidative coupling in the presence of peroxidases and hydrogen peroxide, cross-linking adjacent arabinoxylan (AX) molecules and thus contributing to cell wall assembly (Geissman and Neukom, 1971; Fry et al., 2000; Fry, 2004; Encina and Fry, 2005; Parker et al., 2005; Burr and Fry, 2009). Due to its polysaccharide cross-linking role, phenolic coupling is involved in a number of cell wall properties, causing cell wall stiffening and growth cessation, promoting tissue cohesion, strengthening cell wall structure in response to biotic and abiotic stresses and hindering cell wall degradability (de O Buanafina, 2009).

Occurrence and Functions of Mucilage

A Model of a Mucilage-Supported Diazotrophic Microbiota

Disassembly of the Complex Mucilage Polysaccharide

Sugar Utilization to Support Nitrogenase Activity

Reduction of Oxygen Levels in the Mucilage Environment

Maintaining Low Nitrogen Levels in the Mucilage Environment

Concluding Remarks

Acknowledgement

References

Role for Nitrogen Fixation in Cereal Crops

Nitrogen-fixing microbial associations with nonlegumes, especially cereals, have been a topic of intense interest for more than a century, since such associations could reduce the requirement for nitrogen fertilizers [1–4]. For instance, sugarcane associates with diazotrophic endophytes that contribute to its nitrogen nutrition in some environments [5–7] and a study based on 15N dilution experiments in Miscanthus × giganteus concluded that this bioenergy feedstock could acquire about 16% of its nitrogen from the air [8]. These examples indicate that some monocots have the potential to associate with diazotrophs and acquire small, but significant, amounts of fixed nitrogen from the atmosphere. It has been shown that the model C4 grass Setaria viridis can obtain most of its fixed N from associative nitrogen fixation following inoculation with diazotrophs [9]. The possibility that cereal crops obtain a significant proportion of total N by associative nitrogen fixation has also been suggested by a 50-year assessment of global nitrogen budgets in maize (Zea mays), rice (Oryza sastiva), and wheat (Triticum aestivum), which concluded that up to 24% of the total nitrogen in these crops was derived from nonlegume symbiotic nitrogen fixation [10]. Interestingly, mucilage on aerial roots of sorghum (Sorghum bicolor) with an inset micrograph of Azospirillum brasilense was highlighted on the cover page of the proceedings of a conference on cereal nitrogen fixation held in India in 1984 [11], suggesting that it was suspected more than 35 years ago that sorghum mucilage harbors a diazotrophic microbiota. However, no report on nitrogen fixation in sorghum mucilage was published in the 1984 proceedings volume or elsewhere.

Occurrence and Functions of Mucilage

A key feature of the Sierra Mixe maize landrace mucilage is the abundance of sugars that potentially serve as a source of energy for the diazotrophs. The monosaccharide composition indicated that the mucilage primarily comprises fucose (41%), galactose (36%), arabinose (14%), xylose (3%), glucuronic acid (3%), and mannose (3%) [12]. This composition is similar to the mucilage reported for maize underground roots [15,16]. Osborne et al. found that underground root mucilage contained fucose (61.0%) and glucose (31.4%), while Chaboud found the mucilage to contain galactose (30.7%), fucose (19.3%), glucose (18.5%), xylose (15.2%), and arabinose (13.4%). Both of these previous studies showed high amounts of fucose, xylose, arabinose, and galactose, which agrees qualitatively with the analysis of the Sierra Mixe aerial root mucilage. This monosaccharide composition is not commonly found in plant cell wall polysaccharides and may play a role in signaling associative diazotrophic bacteria that can degrade the mucilage complex polysaccharide and use the released monosaccharides to support growth and nitrogen fixation.

Introduction

The pollution haven hypothesis postulates that dirty industries migrate from high-income countries to low and middle-income countries through the trading of goods and foreign direct investment. Foreign direct investment (FDI) inflows remain one of the main sources of external funding for developing countries, yet, the relocation of carbonintensive and energy-intensive industries from jurisdictions with more stringent environmental regulation to weak locales results in pollution haven. The transfer, dissemination, and diffusion of FDI inflows with polluting technologies, goods, and services to developing countries become the most important part of the challenge to achieve the sustainable development goals (SDGs). On the contrary, the environmental Kuznets curve (EKC) hypothesis postulates that the initial growth of a country’s economic development leads to gradual deterioration of environmental quality and improves environmental conditions after reaching a threshold in economic development (Grossman and Krueger, 1991). Thus, both the pollution haven hypothesis and the EKC hypothesis are important policy derivatives for developing countries. Considering the importance of climate change mitigation and its impacts, as accentuated in SDG 13, the effect of FDI inflows, economic development, and energy consumption on greenhouse gas emissions in developing countries needs further attention to be able to alleviate the impacts. Studies on pollution haven hypothesis (Zakarya et al. (2015) Behera and Dash, 2017; Solarin et al. (2017) Sun et al. (2017), support the validity of this hypothesis. Solarin et al. (2017) validated the pollution haven hypothesis for Ghana using the autoregressive distributed lag (ARDL) bounds testing approach. Sun et al. (2017) examined the impact of FDI inflows, economic growth, energy use, economic freedom, urbanization, financial development, and trade openness on CO2 emissions using the autoregressive distributed lag model. The study confirmed the validity of the pollution haven hypothesis in China and that the positive effect of FDI inflows stems from the large contribution of manufacturing, mining and electricity shifted from the developed countries. Using the fully modified ordinary least squares (FMOLS) and dynamic ordinary least squares regression, Behera and Dash (2017) found a positive impact of FDI inflows and energy consumption on CO2 emissions in 17 south and southeast Asian countries, thus, confirming the pollution haven hypothesis. Zakarya et al. (2015) found a long-run effect of FDI inflows and energy consumption on CO2 emissions in Brazil, Russia, India, and China, thus, validating the pollution haven hypothesis via panel causality and FMOLS regression. On the contrary, studies like Zhu et al. (2016) and Zhang and Zhou (2016), rejected the pollution haven hypothesis. Zhu et al. (2016) employed panel quantile regression to examine the heterogeneous effect of FDI inflows, economic growth, and energy consumption on CO2 emissions in Indonesia, Malaysia, Philippines, Singapore, and Thailand from 1981 to 2011. The study found insufficient support for the pollution haven hypothesis but rather found the halo effect hypothesis in high emission countries. Zhang and Zhou (2016) argue that FDI inflows of modern technologies contribute to CO2 emissions reduction in China rather than environmental deterioration. Dasgupta et al. (1999, 2001) and Dean et al. (2004) revealed that developing countries depend on sophisticated technology transfer through FDI inflows from developed countries as their primary source of acquiring technology. Hence, clean and upgrading from vintage to modern technologies help in the reduction of emission levels.

1- Introduction

2- Theory

3- Material and methods

4- Results and discussion

5- Conclusions

References

Abstarct

During gastric digestion, hydrolysis of proteins by pepsin contributes largely to the breakdown of protein-rich food. We hypothesized that the effect of pepsin is limited by its diffusivity, which is co-determined by the food structure and the local pH in the food during digestion. To investigate the principle mechanism of enzyme diffusion in food matrices, we used enhanced green fluorescent protein (EGFP) as probe to study the diffusivity of proteins in whey protein isolate gels, using fluorescence correlation spectroscopy (FCS). Gels made with different ionic strength showed distinctive elastic moduli but did not show differences in diffusivity of EGFP. Some models for diffusion in hydrogels yield good description of the obtained data, and can approximate the enzyme diffusion in diverse food matrices. However, the enzyme pepsin is more complicated than the probe EGFP, to yield more accurate predictions, electrostatic and enzyme-substrate interaction also need to be considered.

Introduction

The digestion kinetics of food are dependent on the structure of the food that is digested. Food structure influences the oral processing, the gastric disintegration rate, and the consequent gastric emptying towards the duodenum (Singh, Ye, & Ferrua, 2015). The gastric disintegration of food invovles physical and chemical processes, including the peristalsis of the stomach, acid hydrolysis and enzymatic reactions (Bornhorst & Singh, 2014). Among these processes, hydrolysis of proteins by pepsin contributes largely to the breakdown of protein-rich food in the stomach. This hydrolysis can limited by the diffusion of pepsin and the local pH in the solid food matrix during digestion. The hydrolysis kinetics of egg white protein gels and whey protein gels differed strongly from that of the same proteins in solution, which is likely due to the diffusion limitation in gels for both the pepsin and the hydrolysates (Luo, Boom, & Janssen, 2015). Compared to acid-induced dairy gels, a rennet-induced casein gel consists of compact protein aggregates in the acidic gastric environment, and the rennet gel had much slower proteolysis kinetics than that of acid-induced gels (Floury et al., 2018). Thus, a quantitative investigation of pepsin diffusion in food structures may contribute to the understanding of food breakdown and digestion kinetics. We previously measured the diffusivity of pepsin in whey protein isolate (WPI) gels by fluorescence correlation spectroscopy (FCS). We found that the pepsin does not penetrate deep into the gel but remains in a thin layer below the surface of the gel. A second finding was that the diffusivity of pepsin depends strongly on the concentration of the protein gels (Luo, Borst, Westphal, Boom, & Janssen, 2017). Fluorescence Correlation Spectroscopy (FCS) was used for its non-invasiveness and suitability to be used within protein gels. In FCS, a confocal laser microscope is coupled with a photon detector to measure fluorescence intensity fluctuations in a small focal volume. If these fluctuations originate primarily from the diffusion of the fluorophores through the focal volume, autocorrelation analysis can quantify the diffusion rate of the fluorophore. Whey protein gels were used before as model for protein-based solid foods (Luo et al., 2017). Whey protein gelation is generally a two-step process. After heat denaturation, protein oligomers form primary aggregates with different shapes and sizes depending on the pH and the salt concentration. These primary aggregates then form large self-similar aggregates that precipitate or gel above a critical concentration (Aymard et al., 1996; Nicolai, Britten, & Schmitt, 2011).

Recycling ethanol fermentation stillage from food waste could help to resolve the pollution problems, but it would lead to the accumulation of lactic acid (LA), which had a negative effect on ethanol fermentation. In this study, bipolar membrane electrodialysis was used to recover LA as well as other fatty acids in the stillage, and the corresponding parameters that affected this process were investigated. Pretreatment via centrifugation and ultrafiltration reduced the turbidity successfully. Experiments on simulated and real stillage showed that organic acids such as formic acid and acetic acid had a slight effect on the separation of LA, but the existence of chloride ions could form strong anion competition with lactate ions. Simulation experiments with sodium chloride reduced the recovery of LA from 98% to 92%. The recovery rate of LA from real stillage could reach 71.2%. After electrodialysis, the stillage can be reused in ethanol fermentation, the fermentation time was shortened to 24 h, and the ethanol yield increased.

Introduction

For the sustainable development of society, the recycling of waste has become more and more important. Food waste (FW) accounts for a large proportion of municipal solid waste (15%–63%) [1]. It has high organic, volatile solid (85%–95%), and moisture content (75%–85%) and is easily degraded and deteriorated. There are many ways of reusing the waste, such as composting and production of pig feed, biomethane, biodiesel, ethanol, and lactic acid (LA) [2]. The use of FW to produce ethanol can take full advantage of the rich nutrients in FW and obtain fuel. However, a large quantity of stillage will be produced; one way for dealing it is stillage recycle fermentation [3]. This method can reduce pollution and utilize proteins, amino acids, and various metal ions in stillage [4]. In our previous study, LA was determined to be the major accumulated by-product along with stillage recyling. A significant inhibition phenomenon was observed after 5 times recycling. Although the addition of calcium carbonate could alleviate the influence of LA on ethanol fermentation, if we can separate the accumulated LA in the stillage, it will provide better economic benefits. LA is an important natural organic acid. It has received increasing attention as one of the most important building blocks for the production of polylactic acid [5,6]. At present, the main methods of LA production are microbial fermentation and chemical synthesis. Biological fermentation is inexpensive, utilizes a wide range of raw materials, and produces optically pure L- or D-lactic acid, which make it superior to chemical synthesis. More attention has also been paid to the utilization of renewable carbohydrates as a carbon source for LA fermentation [7,8]. However, the major cost of LA production is due to its downstream separation and purification steps [9]. As LA exists in the form of lactate salt in the fermentation broth, acidification and removal of impurities are required for future separation [10]. The extraction methods of LA include precipitation, solution extraction, adsorption, distillation, and nanofiltration [11]. Electrodialysis has the advantages of diversity, sophisticated functions, and technological compatibility [12,13]. Danner et al. used monopolar electrodialysis to extract LA from grass silages and obtained press extraction yields of 31–96 g of lactate per kilogram of silage dry matter (g LA/kg DM) [14]. Chen used conventional electrodialysis (CED) to remove 80% lactate from acid whey [15]. Lopez-Garzon used bipolar membrane electrodialysis (BMED) to recycle LA and other carboxylic acids in the fermentation broth [16].

Production of lactic acid (LA) from lignocellulosic materials has become increasingly prominent in the market. However, for industrial scale to be reached, several challenges in second-generation lactic acid production must be overcome. Besides obstacles such as the hydrolysis process, one of the major challenges of the lactic acid industry is the separation and purification process. This study evaluates the separation of lactic acid produced from hemicellulose hydrolysate from sugarcane bagasse using a Hybrid Short Path Evaporation (HSPE) system. Results showed that a lactic acid concentration of 3.1 times the feed concentration (27.85 g/L) is achievable. Three operational parameters were studied: evaporator temperature, internal condenser temperature and feed flow rate. Maximum lactic acid concentration was 86.69 g/L with an evaporator temperature of 120 °C, condenser temperature of 13 °C, and feed flow rate of 8.27 mL/min. Separation of LA from hemicellulosic sugars using HSPE was a more difficult process than separation of LA from 6-carbon sugars.

Introduction

Lactic acid (LA) is a high added-value product that has been gaining market share every year [1]. It stands out in the environmental, ecological, and medical areas, especially as a building-block molecule. The use of LA has been common in the food industry for a long time and new uses become available every year [2]. LA production by fermentation has several advantages when compared to chemical synthesis, such as low cost of substrates, relatively low required temperatures, low energy consumption, better environmental traits, high purity [3] and easiness to achieve products with tailor-made characteristics [2]. About 90% of all industrially-produced LA worldwide comes from bacterial fermentation [4]. LA biorefineries can exploit various types of feedstock for obtaining second-generation (2G) LA, for example sugarcane bagasse [5], with many advantages such as employment of cheap and non-food competitive raw materials as substrates in the fermentation process. Brazil is the biggest sugarcane producer in the world. The production of sugarcane bagasse in Brazil (2015/2016) has reached 166.4 million tons [6], making this a substrate of great importance in the biofuel and biochemicals scenario. Downstream processes are decisive for LA industry development. This is especially because in medical and pharmaceutical applications, for example, a high-purity product is often required [7]. It is also necessary to consider environmental issues, with increasingly stringent legislation governing the use of solvents and waste generation, in favor of a sustainable production chain [2]. The traditional LA production chain involves a series of downstream treatments like precipitation, conventional filtration, acidification, carbon adsorption, evaporation, crystallization and others. LA separation and purification require a simpler and cheaper alternative to conventional precipitation process, still achieving high purity and yield. In this sense, Short Path Evaporation (SPE) can be an attractive alternative. Economic feasibility analyses of HSPE have been carried out in a few studies, and in any case the proposed method had fewer separation steps in comparison to conventional LA downstream, allowing for a reduction in equipment and operational costs. Furthermore, industrial interests in the purification of LA via SPE is made abundantly clear by the many published patents [8].